4 '-Ethynyl-2'-deoxynucleosides (4'-EdN) represent a new class of nucleoside analogues endowed with potent activity against a wide spectrum of HIV viruses, including some resistant clones. In our laboratory, we have synthesized a 2',3'-dideoxy (4'-EddN) version with cytosine as the nucleobase to reduce toxicity. Although the missing 3'-OH group proved to be critical for biological activity, the 5'-triphosphate of (?)-4'-EddCyt was a potent inhibitor of HIV reverse transcriptase (RT) in vitro. In order to determine which enantiomer was responsible for activity, the nucleoside was resolved and the individual enantiomers converted to the 5'-triphosphate derivatives. Both D- and L- isomers showed inhibitory activity against RT, albeit one enantiomer was clearly more potent than the other. Experiments with a RT (M184V) mutant, which selects against L-enantiomers is underway. Pending identification of the correct enantiomer, a pro-drug that would by-pass the first kinase activation step is planned. We have additionally discovered a new class of anti-HIV agents belonging to the family of conformationally constrained, 2',3'-dideoxybicyclo[3.1.0]hexene analogues which are activated by cellular kinases. Specifically the thymidine analogue, which is conformationally equivalent to the clinically useful agent D4T is a promising agent. This compound showed activity in CEM, MT2 and MT4 cells with little cytotoxicity. Additional studies are in progress. The 5'-triphosphate of the drug inhibited HIV RT with an IC(50) ten-fold higher than that of D4T-5'-triphosphate.